Hedgehog (Hh) signaling is a critical initiator of developmental patterning in many tissues and organs including the heart. Beyond development, Hh signaling acts to promote angiogenesis and epicardial regenerative capacity, preserve function after myocardial infarction, and maintain coronary vasculature in adult myocardium. Cardiac development requires trans-tissue Hedgehog signaling that is specifically received by a subset of cardiac mesodermal cells. This study will test whether Hedgehog is transported by specialized signaling filopodia known as cytonemes from the embryonic ectoderm to its target cells. Cytonemes are thin actin-based filopodia which connect paracrine signal producing cells to receiving cells. Cytonemes have been shown to be the mechanism by which morphogens are dispersed in many other developmental systems but have never been specifically examined in cardiac development. We hypothesize that cytonemes extending between the dorsal ectoderm and cardioblasts are responsible for the specificity of Hh distribution. The presence, function and necessity of Hh- carrying cytonemes for cardiac tube development will be assessed using live confocal imaging of embryos which express fluorescently tagged CD4 and Hh in the Drosophila embryonic ectoderm, mesoderm, or cardioblasts. To further examine how cytonemes distribute Hh, electron microscopy (EM) will be used in the Drosophila larval imaginal wing disc, where cytonemes have been well-established. Because cytonemes share some characteristics of neurons, we will test whether a bouton structure is found at the junction between cytonemes and their target cells. EM imaging will also be used to answer whether cytonemes connect to multiple or single cells, and how Hh is transported along the cytonemes (e.g. membrane associated exosomes, intracellular membrane, endocytic vesicles). Identification of cytoneme-mediated Hh delivery during cardiac development would expand the way in which we understand and potentially target congenital heart defects, and influence future approaches in regenerative medicine.